The proposed studies will focus primarily on supramolecular aspects of biological membranes. Using a combination of freeze-etching, freeze-substitution, negative staining, SDS-polyacrylamide gel electrophoresis and immuno-labeling techniques, we will investigate: a) transient reversible connections between cytoplasmic membranes and plasma membranes; b) the location of specific peptides in spot desmosomes; c) structure-function correlations of chloroplast envelope membrane components. A new propane jet freezing device, which permits a freezing rate of approximately minus 10,000 degrees C/sec will be used to obtain the desired time resolution and structural preservation of the transient membrane phenomena being investigated. The physiological conditions leading to, and the sequence of events associated with the formation of transient connections between cytoplasmic membranes and the plasma membrane will be investigated in fucoid eggs, the red alga Porphyridium, growing tips of pollen tubes, and fungal hyphae. In the desmosome studies, antibodies will be produced against selected polypeptides purified from isolated cow nose spot desmosomes. These antibodies will be used to localize the individual peptides in intact desmosomes. The chloroplast envelope membrane studies will focus on the structural and biochemical characterization of newly discovered differentiated membrane regions.